ABSTRACT Project 1 Recent evidence supports the potential of adeno-associated virus serotype 8 (AAV8)-mediated liver-directed gene therapy as an effective therapeutic intervention for a variety of metabolic disorders. The long-term objective of this project focuses on the clinical development of AAV8 vector-mediated gene transfer for the treatment of two Mendelian disorders of lipid metabolism with limited therapeutic options: homozygous familial hypercholesterolemia (hoFH) and familial lecithin-cholesterol acyltransferase (LCAT) deficiency (FLD). HoFH is caused by loss-of-function mutations in the LDL receptor and characterized by severe hypercholesterolemia, poor response to traditional therapies, and development of severe atherosclerotic cardiovascular disease in childhood or adolescence. FLD is an autosomal recessive condition caused by mutations of the gene encoding for LCAT, an enzyme that esterifies free cholesterol carried by lipoproteins to cholesteryl ester. FLD patients are characterized by markedly low plasma HDL-C and increased plasma FC levels. FLD patients develop chronic renal dysfunction that usually progresses to renal failure by the 4th decade of life. The liver is the primary site of expression of both the LDL receptor and LCAT. Thus, the development of liver-directed gene therapy represents a viable therapeutic approach to both of these diseases. A first generation AAV8 clinical candidate for the treatment of hoFH was identified during the previous cycle of this P01. As a result of the extensive body of work performed, including efficacy and pre-clinical IND-enabling studies, Project 1 is now ideally positioned to initiate clinical studies. In Specific Aim 1, we will perform a phase 1, single ascending dose study in hoFH patients to assess safety of the AAV8-based vector encoding the LDL receptor that has been previously developed by our team. Assessment of safety will include a careful follow-up of laboratory parameters (including immunologic responses) and reported adverse events. In Specific Aim 2, we will measure LDL-C and other lipid parameters before and up to 1 year after the administration of the vector, and conduct lipoprotein kinetic studies before and 3 months after the administration of the vector to directly measure changes in apoB clearance in vivo. The development of an AAV8 vector-mediated gene therapy for FLD will follow a similar approach. Project 2 of the current P01 demonstrated that hepatic expression of human LCAT in FLD mice was able to restore a normal lipid phenotype. In this next cycle, Project 2 will extend this work to optimize the LCAT transgene and establish a mouse model of FLD renal disease to test the effect of AAV8-mediated LCAT gene transfer on renal function. Once a clinical candidate vector is identified by Project 2, we will conduct a study in a murine model of FLD to determine the minimal effective dose of the clinical candidate vector (Specific Aim 3), and the necessary IND-enabling studies in mice and NHP models (Specific Aim 4). Our goal is to be in a position to initiate a clinical trial of an AAV8-based vector encoding LCAT in patients with FLD by the end of the funding cycle.